Recently, a large number of measurements using intermolecular interactions such as immune responses are being carried out in clinical tests, etc. However, since conventional methods require complicated operations or labeling substances, several techniques are used that are capable of detecting the change in the binding amount of a test substance with high sensitivity without using such labeling substances. Examples of such a technique may include a surface plasmon resonance (SPR) measurement technique, a quartz crystal microbalance (QCM) measurement technique, and a measurement technique of using functional surfaces ranging from gold colloid particles to ultra-fine particles. The SPR measurement technique is a method of measuring changes in the refractive index near an organic functional film attached to the metal film of a chip by measuring a peak shift in the wavelength of reflected light, or changes in amounts of reflected light in a certain wavelength, so as to detect adsorption and desorption occurring near the surface. The QCM measurement technique is a technique of detecting adsorbed or desorbed mass at the ng level, using a change in frequency of a crystal due to adsorption or desorption of a substance on gold electrodes of a quartz crystal (device). In addition, the ultra-fine particle surface (nm level) of gold is functionalized, and physiologically active substances are immobilized thereon. Thus, a reaction to recognize specificity among physiologically active substances is carried out, thereby detecting a substance associated with a living organism from sedimentation of gold fine particles or sequences.
In all of the above-described techniques, the surface where a physiologically active substance is immobilized is important. Surface plasmon resonance (SPR), which is most commonly used in this technical field, will be described below as an example.
The surface plasmon resonance method, which enables the measurement of the change in the refractive index on the surface of gold at high sensitivity, is effective for searching a molecule binding to a protein by immobilizing the protein on the gold surface, or for analyzing the binding rate. At the same time, the surface plasmon resonance method is a method for measuring only the change in the refractive index on the gold surface, and thus signals are generated even from insoluble matters (contaminants) contained in a measurement solvent having a refractive index that is different from that of a sample solution. In particular, when contaminants of a large size, such as dusts, exist in a measurement solution, these contaminants cause large noise due to their large volume.
In order to solve this problem, it is conventionally proposed that a sample solution is previously filtrated (Japanese Patent Laid-Open (Kokai) No. 2001-148188). Japanese Patent Laid-Open (Kokai) No. 2001-148188 describes that analysis is carried out in a state where the solution continuously flows. However, such analysis has been problematic in that if the fraction size of a filter used in filtration is small, the filter is easily clogged, and a sufficient amount of solution cannot thereby be collected. In addition, such analysis has been also problematic in that if the fraction size of such a filter is large, contaminants cannot be eliminated to such an extent that it is sufficient for reducing noise. Moreover, although a sufficient amount of the filtrated solution can be obtained by increasing the area of a filter with respect to the amount of a sample solution to be filtrated, the decrease in molecules to be analyzed contained in a sample solution becomes not negligible in this case.
A commonly used measurement chip comprises a transparent substrate (e.g., glass), an evaporated metal film, and a thin film having thereon a functional group capable of immobilizing a physiologically active substance. The measurement chip immobilizes the physiologically active substance on the metal surface via the functional group. A specific binding reaction between the physiological active substance and a test substance is measured, so as to analyze an interaction between biomolecules. Examples of the surface plasmon resonance measurement device for carrying out the analysis as mentioned above include that of Japanese Patent Laid-Open (Kokai) No. 2001-330560.
When a specific binding reaction between a physiologically active substance and a test substance is measured, the binding reaction is generally measured by: connecting in series a reference cell, to which a physiologically active substance interacting with a test substance does not bind, with a detection cell, to which a physiologically active substance interacting with a test substance binds; placing the connected cells in a flow channel system; and feeding a liquid through the reference cell and the detection cell, so as to carry out the measurement of the binding reaction. During the measurement, the liquid contained in the above flow channel system is exchanged from a reference liquid containing no test substance to be measured to a sample liquid containing a test substance to be measured, so as to cause the binding reaction between the physiologically active substance and the test substance to be initiated, and to measure a change in signals due to a lapse of time. However, this measurement method is problematic in terms of the noise width of the change in signals during measurement. Thus, it has been difficult to obtain binding detection data with high reliability.